Methods for the collection of pituitary portal blood have been employed in studies concerned with the detection or measurement of hypothalamic secretion of luteinizing hormone releasing hormone (LHRH) (Fink & Harris, 1970; Harris & Ruf, 1970; Burger et al., 1972; Fink & Jamieson, 1973, 1974; Carmel, 1975; Eskay et al., 1975; Ching, 1976) and thyrotropin releasing hormone (TRH) (Averill et al., 1966; Wilbur & Porter, 1970; Ching & Utiger, 1976) in rats. Up until now, the method of portal blood collection has been adequate for the detection of these neurohormones in portal blood. However, the validity of its use for the measurement of hypothalamic secretion of LHRH and TRH has not been established. As a consequence, the present study is designed to determine whether or not measurement of LHRH and TRH in portal blood, collected in a similar manner to that reported by Porter and Smith (1967), constitutes a reliable means of assessing hypothalamic secretion of these releasing hormones. In this regard, the aims of this study are to examine the effects of several important parameters that could influence our ability to assess accurately the hypothalamic release of LHRH and TRH. These are: (1) the physiological state of the donor rat, (2) the protocol for the collection of portal blood, (3) recovery of LHRH and TRH from blood for assay, (4) time-related distribution of LHRH and TRH in brain and blood, and (5) method of measuring LHRH and TRH in blood. Following the establishment of the most reliable and valid procedure for the collection of portal blood and quantification of LHRH and TRH therein, physiological studies involving gonadectomy and thyroidectomy, and replacement therapy by the appropriate target gland hormone are to be enacted to further test the methodologies and experimental protocols and elucidate the manner by which target gland hormones affect hypothalamic release of the appropriate releasing hormone.